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The endonuclease activity of Mili fuels piRNA amplification that silences LINE1 elements

Serena De Fazio1, Nenad Bartonicek, Monica Di Giacomo

  • 1European Molecular Biology Laboratory, Mouse Biology Unit, Via Ramarini 32, Monterotondo Scalo 00015, Italy.

Nature
|October 25, 2011
PubMed

Insights

Piwi proteins like Mili are crucial for amplifying piRNAs (Piwi-interacting RNAs) to silence LINE1 transposons. Disrupting Mili

Area of Science:

  • Reproductive Biology
  • Molecular Genetics
  • Epigenetics

Background:

  • Piwi proteins and piRNAs are essential for transposon silencing, particularly during germline development.
  • Mili and Miwi2 are murine Piwi proteins involved in epigenetic silencing of LINE1 and intracisternal A particle (IAP) transposons.
  • The precise role of Piwi proteins' endonuclease activity in piRNA biogenesis and transposon control remains unclear.

Purpose of the Study:

  • To investigate the functional significance of Piwi-catalyzed endonuclease activity in piRNA biogenesis and transposon silencing.
  • To elucidate the specific roles of Mili and Miwi2 in these processes by analyzing catalytically inactive mutants.
  • To determine the contribution of Piwi endonuclease activity to LINE1 and IAP transposon silencing and male fertility.

Main Methods:

  • Generation of mice with point mutations in the catalytic DDH triad of Mili and Miwi2, creating Mili(DAH) and Miwi2(DAH) alleles.
  • Analysis of piRNA populations and transposon silencing in gonadocytes from mutant mice.
  • Assessment of fertility and spermatogenesis in Mili(DAH) and Miwi2(DAH) homozygous mice.

Main Results:

  • Mili(DAH) mutation disrupted piRNA amplification, leading to reduced piRNAs in Miwi2 complexes and impaired LINE1 silencing.
  • Mili-mediated piRNA amplification is essential for LINE1 silencing but not for IAP silencing.
  • Homozygous Miwi2(DAH) mice were fertile, with normal transposon silencing and secondary piRNA biogenesis, indicating Mili's primary role in amplification.
  • Defective piRNA pathway in Mili(DAH) mice caused spermatogenic failure and sterility.

Conclusions:

  • Intra-Mili secondary piRNA biogenesis drives piRNA amplification, which is indispensable for LINE1 silencing.
  • Mili's endonuclease activity is critical for piRNA amplification and LINE1 silencing, impacting male fertility.
  • Miwi2's endonuclease activity is not essential for piRNA amplification or LINE1 silencing in the male germline.

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